A Review of the Efficacy of Ultraviolet C Irradiation for Decontamination of Pathogenic and Spoilage Microorganisms in Fruit Juices

Ultraviolet C (UV-C, 200–280 nm) light has germicidal properties that inactivate a wide range of pathogenic and spoilage microorganisms. UV-C has been extensively studied as an alternative to thermal decontamination of fruit juices. Recent studies suggest that the efficacy of UV-C irradiation in reducing microorganisms in fruit juices is greatly dependent on the characteristics of the target microorganisms, juice matrices, and parameters of the UV-C treatment procedure, such as equipment and processing. Based on evidence from recent studies, this review describes how the characteristics of target microorganisms (e.g., type of microorganism/strain, acid adaptation, physiological states, single/composite inoculum, spore, etc.) and fruit juice matrices (e.g., UV absorbance, UV transmittance, turbidity, soluble solid content, pH, color, etc.) affect the efficacy of UV-C. We also discuss the influences on UV-C treatment efficacy of parameters, including UV-C light source, reactor conditions (e.g., continuous/batch, size, thickness, volume, diameter, outer case, configuration/arrangement), pumping/flow system conditions (e.g., sample flow rate and pattern, sample residence time, number of cycles), homogenization conditions (e.g., continuous flow/recirculation, stirring, mixing), and cleaning capability of the reactor. The collective facts indicate the immense potential of UV-C irradiation in the fruit juice industry. Existing drawbacks need to be addressed in future studies before the technique is applicable at the industrial scale.

forming units (CFU)/ml [30]. In another study, batch UV-C treatments (11.50 or 13.44 W/m 2 for 8 min, and 0.34 W/m 2 for 25 min) produced 5 log reductions of A. acidoterrestris spores in apple juice [31]. For the inactivation of fungal spores, application of a batch UV-C treatment (36 W/m 2 ) for 10 min allowed the 5.7 log reduction of Aspergillus fischeri and 4.2 log reduction of Paecilomyces niveus in apple juice. When batch UV-C irradiation was applied at a much lower intensity of 6.5 W/m 2 , such reductions were not achieved, even after 30 min of exposure [32]. Inactivation of ascospores of Talaromyces macrosporus and Neosartorya spinosa in apple juice treated with batch UV-C treatments (doses of 7.2, 14.3, or 21.5 J/ml for 1-3 cycles) was reported. However, most of the tested UV-C treatments were not sufficient to attain 5 log reductions of the ascospores, except for N. spinosa ascospores treated with 21.5 J/ml UV-C [33].

Efficacy of UV-C Varies depending on Juice Matrices
Optical and physical characteristics of the treated food matrix, such as UV absorbance (UVA), UV transmittance (UVT), turbidity (nephelometric turbidity units, NTU), soluble solids content (°Brix), pH, and color, influence the efficacy of UV-C treatment. Generally, UV-C efficacy will be lower for a more complex food matrix (e.g., juices that are turbid, colored, and/or suspended solids) [16,29,34]. Evidence from recent studies ( Table 1) clearly shows that the juice matrix has a remarkable effect on UV-C efficacy. Thus, the use of the same UV-C treatment for the decontamination of different types of juices may result in different reductions of specific target microorganisms. For instance, a pilot-scale continuous UV-C treatment (0.39 J/cm 2 ) was highly effective for decontamination of clear pear juice (UVT 89.1%, UVA 0.05/cm, 21.9 NTU), achieving up to 4.4 and 5.5 log reductions for L. plantarum and E. coli, respectively [21]. However, a much lower efficacy of the same UV-C treatment was observed after decontamination of turbid juice blends, yielding less than 4 log reductions of both bacteria in both blended OT (UVT 20.9%, UVA 0.68/cm, 3,100 NTU) and OBMKS (UVT 42.6%, UVA 0.37/cm, 1,767 NTU) juices. Overall, the results of this study suggest that the higher values of UVT and lower values of UV absorbance, turbidity, and color parameter (a*) of the juice samples are closely associated with the higher efficacy of the tested UV-C treatment [21]. Turbid juices with suspended solids have a higher UV absorbance than clear juices. An increase in the level of UV absorbance leads to a decrease in the ability of UV-C light to penetrate food, thereby reducing the antimicrobial efficiency of any UV-C dose [35]. Moreover, the presence of suspended solids and soluble components in the food matrix can weaken the effects of UV-C irradiation by inducing light scattering, absorption, and reflection [15]. In addition, differences in the pH of juice samples are also likely to affect the efficacy of UV-C applied at 1.5 mW/cm 2 for up to 12 min in a batch reactor, in which S. Typhimurium cells from all three growth phases exhibited less reduction in apple juice with an adjusted pH of 5.65, compared to apple juice with a lower pH of 3.63 [26]. Therefore, the development and application of UV-C irradiation for microbial inactivation should encompass a broad range of juice products.

Efficacy of UV-C Varies depending on UV-C Decontamination Procedure
The comparison of UV-C decontamination procedures from different studies is challenging because the results have varied in terms of scale (e.g., pilot-scale vs lab/small scale) and equipment and processing parameters (see Table 1 for more details). Moreover, most studies only reported the applied and/or incident UV-C doses/ intensities, even though the absorbed dose (the irradiant energy absorbed by the food components available for driving the solution reaction) and delivered dose (the actual amount of irradiant energy delivered to the microorganism) are more critical for enhanced microbial inactivation [29]. In general, the results from most of the studies (Table 1) reveal that higher UV-C doses/intensities led to higher rates of microbial inactivation, regardless of the target microorganism and juice sample characteristics. To ensure that the dose/intensity from the UV-C light source is reliably delivered to the irradiated product and enhances the antimicrobial efficiency, a specially designed UV-C continuous or batch reactor with different equipment and processing parameters is necessary. Numerous UV-C continuous reactors, including curved and coiled tubes, and batch reactors, including petri dish, well plate, and tank, with different arrangements and configurations have been developed and applied for the decontamination of fruit juices ( Table 1).
The main part of the UV-C reactor is the UV-C lamp/light source. The target microorganisms in the juice samples are exposed to UV-C light at a certain intensity/irradiance for a certain time. Therefore, selecting a UV-C lamp with appropriate features for microbial inactivation, which include wavelength, power, and size, is an important step to enhance the penetration of UV-C light into juice samples. The efficiency of continuous and batch UV-C decontamination procedures for microbial inactivation can be greatly affected by the UV-C lamp features (e.g., type, number, and position of the lamp and its outer case/layer) and irradiation time. In recent years, different types of UV-C germicidal lamps with various wavelengths ranging from 200 to 280 nm, such as kryptonchlorine excimer lamps (222 nm), general UV-C lamps (253-254 nm), and light emitting diode lamps (254-279 nm), have been studied for their antimicrobial efficacy ( Table 1). The effects of using one or two UV-C lamps (30 or 80 cm length) connected in series on the reduction of S. cerevisiae in grape juice have been reported. No significant difference in the obtained log reductions (approximately 2 log CFU/ml) was observed between the 30 and 80 cm UV-C lamp. However, by increasing the number of tested UV-C lamps, the inactivation efficacy was enhanced (>5 log CFU/ml) because the UV-C intensity was doubled [36]. The results summarized in Table 1 also show that an increase in the irradiation time would increase the UV-C dose/intensity, allowing greater reductions in the target microorganisms in fruit juice samples [19,25,26,28,33,[37][38][39][40]. However, in some cases, further irradiation up to certain exposure times did not cause any significant increase in the microbial reduction [24,32]. Furthermore, several equipment and processing parameters applied in the UV-C decontamination procedure    (Table 1), such as reaction tube/tank/plate (e.g., size, thickness, volume, diameter, outer case, configuration/ arrangement), pumping/flow system (e.g., sample flow rate and pattern, sample residence time, number of cycles), homogenization (e.g., continuous flow/recirculation, stirring, mixing), and cleaning capability of the reactor, may also influence the efficacy of the UV-C decontamination process. All of these parameters need to be evaluated during the development and optimization of UV-C decontamination procedures. UV-C light treatment is more frequently used for surface sterilization because it does not penetrate food samples very deeply. One of the strategies that can be applied for juice treatment is the use of a thin layer to increase the surface area and decrease the depth of the product [13]. For instance, using a reaction tube with a lower thickness (0.5 cm) led to enhanced reductions in S. cerevisiae [36]. Reducing the thickness of the juice film was highly effective, because greater reductions in S. cerevisiae were achieved at any of the tested flow rates, even when using a single UV-C lamp. The results may reflect the narrower distance between the juice sample and UV-C lamp in thinner reaction tubes, thus permitting better light penetration and yeast inactivation [36]. The higher log reductions of E. coli and L. monocytogenes in tender coconut water conveyed in a thinner reaction tube (inner diameter of 1.6 mm vs 3.2 mm) and treated with continuous UV-C irradiation were also reported [20]. However, reducing the thickness of the tube/juice film might cause laminar flow when the juice matrix passes through the reactor. Consequently, UV-C light mostly penetrates the outer layers of the flowing liquid, leaving the inner layers of the flowing liquid less exposed, which could restrict the delivery of a uniform dose/intensity into the sample [33].
To increase the probability of UV-C light penetrating the sample and delivering uniform dose/intensity, the juice solution in the UV-C batch reactor can be mixed using a magnetic stir bar [40] or agitation system for the solution treated in the tank [18]. In the UV-C continuous reactor, the proper mixing of juice solutions can be estimated using the Reynolds and Dean numbers, which indicate the presence of turbulence and secondary flow, respectively, inside the UV-C reactor for any liquid food [41]. The presence of turbulent and secondary flows reportedly allows better mixing of the juice matrix inside the continuous reactor and higher exposure to UV-C light [20,33]. Moreover, to allow for additional mixing, the samples were recirculated or cycled multiple times through the system. Recirculation of the juice solution compensates for the lack of turbulent flow and increases the probability that more parts of the juice matrix can be exposed to UV-C light. Increasing number of cycles increase the greater of UV-C exposure, and consequently increases the effectiveness of the treatment even at the same dose [33]. Multiple cycles of UV-C irradiation were more effective than single cycle for the inactivation of bacteria, yeast, and mold in various fruit juices [21][22][23]33].
Since the UV-C dose is directly proportional to the average residence time and inversely proportional to the flow rate [19], enhanced microbial inactivation might also be achieved by controlling the flow rate or increasing the residence time to increase the delivered UV-C dose. For instance, in one study a lower flow rate of the sample  led to a higher residence time of the sample, greater delivered dose, and increased efficacy of continuous UV-C treatment on the reduction of E. coli and S. cerevisiae in pomegranate juice [23]. However, in another study, an increase in the tested flow rates (from 5.2 ml/s to 17.1 or 31 ml/s) reportedly increased the reduction of S. cerevisiae in grape juice continuously treated with UV-C for 60 min [36]. This discrepancy might be due to other processing factors, such as the features of reaction tubes (e.g., size, thickness, volume, diameter, outer case, configuration/arrangement) and flow pattern (laminar or turbulent flow) that affect the uniformity of the dose delivered to the juice sample and target microorganisms.
Cleaning the UV-C reactor before and after use is necessary to maintain cleanliness and sterility. Several methods that have been used include cleaning with 500 ml of hot water (70°C) followed by 100 ml of hypochlorite (200 ppm) for 10 min, and sterile deionized water at room temperature for 4 min [20]. In other studies, cleaning by recirculating sterile water (~200ml/min) at room temperature for 3 min [23] or sequential flushing with sterile water, 0.1 N HCl, sterile water, 0.1 N NaOH, and a final rinse with sterile water [19] have also been performed. No remarkable changes in the pH, °Brix, acidity, and color parameters (UV-C up to 1.42 J/cm 2 ), but TPC significantly decreased (UV-C at ≥0.28 J/cm 2 ) [46] Grape juice UV-C (254 nm; 10-11 mW/cm 2 for 60 min)

Table 2. Effects of ultraviolet C (UV-C) irradiation treatments on quality properties of different fruit juices.
No significant changes in the °Brix and pH were observed. Significant changes in the color parameters (L, a , b) were observed, but they are not perceptible by the human eye [36] UV-C (254 nm; 19.7 mW/cm 2 for ≥20 min) No significant changes were observed in the pH, acidity, °Brix, turbidity, and color. Significant loss of vitamin C (92%), TPC (19%), and TAA (54%) was observed [40] Carrot juice UV-C (253.7 nm for 5 cycles; dose per cycle 1,152 J/L or 0.23 J/cm 2 ) No significant changes in the physicochemical (color, browning index, viscosity, optical density, density, pH, and turbidity) and sensory parameters were observed [48] Tomato UV-C alone: TPC was not affected but vitamin C and anthocyanin reduced. UV-C combined with ChNPs (100 μl/ ml): TPC, vitamin C, and anthocyanin were not affected [42] Carrotorange juice UV-C (253.7 nm for 15 min; total dose 10.6 kJ/m 2 ) combined with/ out mild heat (50°C) No significant changes in the pH, °Brix, and turbidity were observed. Changes in the TPC, TAA, and color were observed in the UVC-treated samples [49] Orangetangerine juice UV-C (253.7 nm for 0-15 min; total dose 0-1.72 J/cm 2 ) No significant changes were observed in the color, pH, acidity, °Brix, and turbidity (UV-C up to 1.72 J/cm 2 ). TPC and TAA (DPPH) decreased significantly (UV-C at 1.72 J/cm 2 )

Effects of UV-C Irradiation on Quality of Fruit Juices
The effects of UVC irradiation on the quality attributes of fruit juices should be evaluated when developing and applying this treatment. As shown in Table 2, most studies have reported no significant changes in the physicochemical properties that include °Brix, viscosity, optical density, density, pH, acidity, turbidity, browning index, reducing sugar, and color of fruit juices after UV-C irradiation. However, in some cases, adverse effects of UV-C irradiation on the bioactive compounds, including phenolics, antioxidants, anthocyanin, and vitamin C, and color of the treated fruit juices were observed.

Conclusions and Future Perspectives
Proper control of pathogenic and spoilage microorganisms during the processing of fruit juices is a prerequisite for ensuring food safety and maintaining hygiene standards. Based on evidence from recent studies, UV-C irradiation has shown immense potential in the fruit juice industry, and can be a good alternative to traditional thermal decontamination techniques. However, certain drawbacks restrict their industrial scale application. For instance, the microbial inactivation efficacy of UV-C irradiation is largely affected by the characteristics of target microorganisms and juice matrices. The low penetration of UV-C light demands higher doses and/or longer exposure times for the inactivation of highly resistant microorganisms, including yeasts, molds, and spore forming microorganisms, and decontamination of highly complex juice matrices (e.g., turbid, large area/volume, colored). Such prolonged exposure to high UV-C doses may negatively impact the quality of juice, exceed the standards/permissible limits set by regulatory authorities, and may not be industrially feasible. Optimizing the UV-C treatment procedure, for example by using a thin layer reaction tube, may prevent light penetration losses, ensure a uniform delivered dose with shorter treatment time, and enhance the efficacy. However, it can be a handicap when scaling up to the industrial scale, in which the surface-to-depth ratios or flow rate should be maximized. Therefore, for UV-C irradiation technology to be accepted and effectively transferred to the juice industry, future studies should focus on process optimization/validation by taking into account the main factors (target microorganisms, juice matrices, and important parameters of UV-C treatment procedure) that affect treatment efficacy. The responses evaluated for such process optimization/validation should focus on microbial reduction and also on the quality (physicochemical, sensorial, and nutritional attributes) of fruit juices. A combination of UV-C irradiation with other food processing/preservation techniques can also be considered for enhanced efficiency and better outcomes. In addition, studies should be carefully performed at a lab/small scale before its realization at an industrial scale using the available/newly developed industrial scale UV-C decontamination device.